The mitochondrial DNA is 16 times more prone to oxidative damage and evolves 10-20 times faster than nuclear DNA. Point mutations and deletions in this mitochondrial genome give rise to a wide range of diseases, such as Leber's hereditary optic neuropathy, retinitis pigmentosa, Kearn-Sayre syndrome, Pearson marrow pancreas syndrome and ocular myopathy. These mutations may occur during replication by the DNA polymerase gamma. The DNA polymerase gamma differs form the nuclear DNA polymerases due to its sensitivity to antiviral nucleotide analogs, such as AZT and dideoxynucleotides. Patients treated with AZT develop a ragged-red fiber myopathy associated with a reduction in mitochondrial DNA levels. How the mitochondrial DNA polymerase makes point and deletion mutations and what structural properties set this polymerase apart from the nuclear DNA polymerases to give rise to its inhibition patterns are poorly understood. To address these questions we have cloned the DNA polymerase gamma genes and cDNA from S. pombe, D. melanogaster and Homo Sapiens. The recombinant human mitochondrial DNA polymerase gamma protein has been functionally overexpressed greater than 100 fold in insect cells by a recombinant baculovirus and in E. coli. The overexpressed protein is currently being purified to homogeneity and enzymatically characterized. Two mutant DNA polymerase gamma proteins were made by site-specific mutageneisis; an exonuclease deficient and a dideoxy-resistent DNA polymerase. Antibodies against the human pol gamma were used to study the regulation of polymerase gamma expression in human cells in the presence and absence of mitochondrial DNA in human cells. DNA polymerase gamma protein and mRNA levels in mitochondrial deficient cells were the same as their parental cells, suggesting no feedback control from the mitochondria to the nucleus.